Project Summary/Abstract Immune-mediated sensorineural hearing loss (IMSNHL) is of clinical importance because of progressive bilateral involvement and potential reversibility. Currently, the therapeutic targets specific to IMSNHL are not available because its molecular pathogenesis is poorly understood. Our long-term goal is to elucidate the molecular mechanism of immune-mediated inner ear damage, resulting in SNHL. Etiopathogenesis of IMSNHL is largely unknown, but regardless of the initiating factors, the final course of IMSNHL is commonly associated with immune-mediated damage of the cochlear cells (sensory and non- sensory). Immune-mediated tissue damage generally involves direct cell-cell signaling via surface molecules such as antibody-dependent cytotoxicity; and indirect signaling via cytokines such as tumor necrosis factor-a (TNF-a). Previously, we have demonstrated that spiral ligament fibrocytes (SLFs) release mediators in response to inflammatory signals and SLF-derived molecules directly attract the inflammatory cells. Therefore, we hypothesize that SLFs are key players in immune-mediated inner ear damage by responding to inflammatory mediators and releasing chemoattractive and cytotoxic molecules. We also found that secretion of TNF-a is required for cisplatin-induced ototoxicity and that down-regulation of TNF-a attenuates cisplatin-induced auditory damage. These results led us to focus on TNF-a-mediated cytotoxicity as a mechanism of immune- mediated inner ear damage. However, the auditory sensory cells appeared to be damaged only by the extremely high concentration of TNF-a in animal experiments, indicating the requirement of sensitization for TNF-a-mediated inner ear cytotoxicity. Therefore, we aim to determine the molecular mechanism involved in: (1) SLF's TNF-a induction in response to IL-1, a model inflammatory mediator; and (2) sensitization of the auditory sensory cells to induce TNF-a-mediated cytotoxicity via interferon-g (IFN-g). In addition, we plan to determine the therapeutic potential of targeting the key molecules involved in IFN-g-sensitized TNF-a-mediated cytotoxicity, which will provide us with a scientific basis for the development of a novel strategy to manage IMSNHL.